Protein palmitoyl transferase DHHC16 and application thereof in improving salt tolerance of rice
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A rice and protein technology, applied in the biological field, can solve problems such as threats to the safe production of rice
Pending Publication Date: 2022-03-18
HUNAN HYBRID RICE RES CENT
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However, in rice production, there are often stresses from abiotic factors, such as drought, salt damage, high temperature, low temperature...
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Embodiment 1
[0086] Embodiment 1, the cloning of rice protein palmitoyl transferase DHHC16
[0087] The cDNA of rice 9522 was used as a template, and DH16-FL-F (5'-ATGGAGCAGCAGCCGCTGCT-3') and DH16-FL-R (5'-CTAAAGAAGTTTTGAGCTCG-3') were used as primers for PCR amplification. The amplified product is a DNA fragment with a size of about 1000bp. After sequencing and identification, the DNA fragment is 1050bp in length, which is the DHHC16 gene. In the sequence list, SEQ ID No.2, the DHHC16 gene encodes a protein palmitoyl transferase DHHC16 consisting of 349 amino acid residues. For the amino acid sequence of DHHC16, see SEQ ID No. 1. In rice 9522, the genomic DNA of the DHHC16 gene is 4025bp in full length, and its sequence is shown in SEQ ID No.3, which includes 4 exons, namely, the 183-208th, 1341-1575th, 1341-1575th, and No. 1697-1993, No. 2093-2584.
Embodiment 2
[0088] Example 2, Selection of rice DHHC16 target site and construction of site-directed editing vector
[0089] In this example, the rice DHHC16 gene was edited through the CRISPR-Cas9 system, and it was found that the DHHC16 gene and its encoded protein DHHC16 can regulate the salt tolerance of rice. The steps are as follows:
[0090] 1. Selection of target site sequence
[0091] The 1911-1930 positions of the genomic DNA of the DHHC16 gene (ie, the 1911-1930 positions of SEQ ID No. 3) were selected as the target sequence of the CRISPR-Cas9 system, which was located on the third exon of the DHHC16 gene.
[0092] 2. Construction of fixed-point editing recombination vector
[0093] 1. Construction of intermediate vector pYLgRNA-U3-DHHC16
[0094] (1) Design and synthesis of DHHC16 target site linker primers
[0095] Add GGCA before the 5' end of the sense strand of the target sequence, and add AAAC before the 5' end of the antisense strand to obtain the target site adapter ...
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Abstract
The invention discloses a protein palmitoyl transferase DHHC16 and an application thereof in improving the salt tolerance of rice. The DHHC16 disclosed by the invention is A1), A2) or A3) as follows: A1) a protein of which the amino acid sequence is SEQ ID No.1; a2) a protein which is obtained by substitution and/or deletion and/or addition of one or more amino acid residues on the amino acid sequence as shown in SEQ ID No.1 in the sequence table and has the same function; and A3) a protein which has 90% or more of identity with the protein in A1), is derived from rice and has the same function. The homozygous mutant of the DHHC16 gene is subjected to high-salt stress treatment, and it is found that the salt tolerance of the plant is improved due to function deficiency of the DHHC16. Therefore, the DHHC16 and the coding gene thereof disclosed by the invention can be used for regulating and controlling the salt tolerance of the plant and have a good application prospect.
Description
technical field [0001] The invention relates to a protein palmitoyl transferase DHHC16 and its application in improving rice salt tolerance in the field of biotechnology. Background technique [0002] Rice (Oryza sativa L.) is one of the most important food crops in my country and even in the world. Nearly half of the world's population uses rice as a staple food, and the planting area accounts for about one-third of the total area of food crops. However, in rice production, there are often stresses from abiotic factors, such as drought, salt damage, high temperature, low temperature, waterlogging, heavy metals and ozone, which seriously affect the growth and development of rice and threaten the safety of rice production. Therefore, digging out more key genes involved in abiotic stress, exploring more molecular mechanisms of abiotic stress response, and breeding more breeding materials resistant to abiotic stress will be an urgent problem for researchers and breeders. The...
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